We will investigate the regulation of tyrosine synthesis in rat hepatoma and normal rat liver cells in culture. The long range goals are to understand qualitatively and quantitativley the degradation and synthesis of essential metabolites, understand the mechansims through which control of these processes is exerted, and establish in what ways the tumor and normal cell populations differ. Metabolic control (involving effects on substrate and product concentrations) and gentic control (involving effects on mRNA and protein synthesis and degradation) of the rate of hydroxylation of phenylalanine to tyrosine will be examined. We will examine in particular detail on pure phenylalanine hydroxylase, in the liver tumor cells, and in the normal liver cells, the regulation of phenylalanine hydroxylase by its substrate phenylalanine, its cofactor tetrahydrobiopterin, and by iron (which is required for the activity of the enzyme). The mechanisms of action of glucocorticosteroids and the hormones insulin and glucagon on phenylalanine hydroxylase expression in the hepatoma cells and normal liver cells (primary hepatocytes) will be investigated. In these studies, we will relate the concentrations of enzymes, substrates, and cofactors to the functional rate of tyrosine formation by the two cell types. These studies apply biochemical methods to a basic problem of normal liver and liver tumor cell metabolism and to studies of the biology and control of a complete enzyme system. This system is required for normal animal development and its deficiency, as in the human disease phenylketonuria, has serious consequences. It, furthermore, provides a good model system for understanding the regulation of the metabolism of all essential metabolites.